ORL 2013;75:46–53

DOI: 10.1159/000347083 © 2013 S. Karger AG, Basel

Received: October 25, 2012 0301–1569/13/0751–0046$38.00/0
Accepted after revision: January 11, 2013 www.karger.com/orl
Published online: May 9, 2013

Original Paper

Freely-Available, True-Color Volume Rendering

Software and Cryohistology Data Sets for Virtual
Exploration of the Temporal Bone Anatomy
Lüder Alexander Kahrs Robert Frederick Labadie
Department of Otolaryngology, Head and Neck Surgery, Vanderbilt University Medical Center,
Nashville, Tenn., USA

Key Words
Histology · Temporal bone anatomy · Three-dimensional viewer

Abstract
Background: Cadaveric dissection of temporal bone anatomy is not always possible or feasible
in certain educational environments. Volume rendering using CT and/or MRI helps under-
standing spatial relationships, but they suffer in nonrealistic depictions especially regarding
color of anatomical structures. Freely available, nonstained histological data sets and software
which are able to render such data sets in realistic color could overcome this limitation and be
a very effective teaching tool. Methods: With recent availability of specialized public-domain
software, volume rendering of true-color, histological data sets is now possible. We present
both feasibility as well as step-by-step instructions to allow processing of publicly available
data sets (Visible Female Human and Visible Ear) into easily navigable 3-dimensional models
using free software. Results: Example renderings are shown to demonstrate the utility of these
free methods in virtual exploration of the complex anatomy of the temporal bone. After ex-
ploring the data sets, the Visible Ear appears more natural than the Visible Human. Conclusion:
We provide directions for an easy-to-use, open-source software in conjunction with freely
available histological data sets. This work facilitates self-education of spatial relationships of
anatomical structures inside the human temporal bone as well as it allows exploration of sur-
gical approaches prior to cadaveric testing and/or clinical implementation.
Copyright © 2013 S. Karger AG, Basel

Background

Teaching anatomy and learning safe surgical approaches to the temporal bone are chal-
lenging because of the high density of critical anatomical structures embedded within. Espe-
cially difficult to appreciate are the narrow access paths and small dimensions required to

Lüder Alexander Kahrs

Leibniz University Hannover
Institute of Mechatronic Systems
Appelstrasse 11a, 30167 Hannover (Germany)
E-Mail lueder.kahrs @ imes.uni-hannover.de
 ORL 2013;75:46–53 47
DOI: 10.1159/000347083 © 2013 S. Karger AG, Basel
www.karger.com/orl
Kahrs et al.: Freely-Available, True-Color Volume-Rendering Software and
Cryohistology Data Sets for Virtual Exploration of the Temporal Bone Anatomy

reach the middle and inner ear for surgical interventions. This necessitates years of training,
strong experience, and an innate sense of spatial relationships. During medical education,
hand drawings, videos, enhanced photographs from microscopes or endoscopes, and imaging
(e.g. CT or MRI) are used to help discern 3-dimensional (3D) relationships from 2-dimen-
sional images [1, 2]. Complementing this is specialized training for surgical residents during
temporal bone laboratory courses using cadaveric bones as well as hands-on learning under
the close supervision of an expert as is only achievable in the operating room [3, 4]. These
courses are considered the penultimate training second only to apprenticing within the oper-
ating room. Unfortunately, especially in the UK, such courses are in decline secondary to
decreased availability of donated bodies [3].
Beyond standard biomedical image computing software [5], several specific computer
systems have been developed to enhance teaching and understand temporal bone anatomy
[4, 6, 7]. These systems provide virtual simulations [8, 9] some of which are enhanced with
haptic feedback [8, 10–13]. Different visualization methods and applications allow rendering
of full-color image stacks such that they appear in life-like color [14–20], but there are very
few which are available in the public domain and are open source [21, 22]. Despite this, input
data for true-color rendering histology slices (rendering of full-color images) are freely
available from whole body human cryohistology [23–25]. More detailed data sets of the
subregion of the ear/temporal bone are also available [26–28]. The aim of this work is to give
instructions for a free method for visual exploration of otological anatomy with 3D, true-color
volume rendering using these data sets.

Methods

After a critical analysis of visualization software available in the public domain, we selected the advanced
volume rendering technique, introduced by Peng et al. [29, 30] formerly referred to as V3D (Viatronix Inc.,
Stony Brook, N.Y., USA) now freely available, including source code for nonprofit research, under the name
Vaa3D. The main advantage of Vaa3D over other software is the true-color rendering capability combined with
real-time interactive visualization (i.e. no render button is required and views are immediately updated after
changing the view). The software runs on typical PCs and laptops with the requisite memory and a dedicated
graphics card. Our test environment was a standard personal computer (laptop Lenovo Y470) with the following
specifications: Intel i7 processor (2630QM, 2 GHz), 8 GB RAM and 1 GB dedicated graphics card memory (Nvidia
GeForce GT550M) available for under USD 1,000. Software versions are available for download on the Vaa3D
website (www.vaa3d.org). We downloaded Vaa3D version 2.707 for Windows 7 (64 bit). For comfortable visu-
alization we recommend a display which can provide an application window height of at least 768 pixels.
Immediately after downloading the Vaa3D software is ready to use. Image series – i.e. multiple TIFF
(tagged image file format) images – can be imported to an image stack, or image stacks (e.g. multi-page TIFF
files) can be loaded directly. We tested two different data sets which include or consist of the temporal bone
region: (1) the Visible Female Human [31] and (2) the Visible Ear [26]. Both data sets are nonstained histo-
logical slices and were originally acquired by cryosectioning and color photography.
(1) The Visible Human data sets are public domain (written permission is necessary prior to use) and
can be downloaded via FTP from the US National Library of Medicine after receiving the necessary access
data. We downloaded the ‘4K-Tiff-Images’ of the Visible Female Human and used image file numbers 1271–
1500 which cover the temporal bone. Using freeware (Irfanview 4.32, www.irfanview.com, free for noncom-
mercial use) we cropped these for a left and a right ear region (fig. 1a). Both resulting image stacks have 512
× 512 × 230 voxels, and the in-plane resolution is higher (×2.4) than in the z-direction (slice distance 0.33
mm). Such anisotropy (and noninteger/non-whole-number) scaling factors need to be entered in Vaa3D. The
resulting volumes cover regions of 7.04 × 7.04 × 7.59 cm3.
(2) The Visible Ear data set, created by Sørensen et al. [26], is published as freeware, and a CD can be
ordered from Karger AG, Basel, Switzerland. The files on the CD can be copied for noncommercial research
and educational purposes. The data set is comprised of 605 slices of a human temporal bone, each with an
image dimension of 3,078 × 1,942 pixels and in-plane resolution of 50 μm/pixel. As shown in figure 1b, we
 ORL 2013;75:46–53 48
DOI: 10.1159/000347083 © 2013 S. Karger AG, Basel
www.karger.com/orl
Kahrs et al.: Freely-Available, True-Color Volume-Rendering Software and
Cryohistology Data Sets for Virtual Exploration of the Temporal Bone Anatomy

Color version available online

a b

Fig. 1. Exemplary slices of each data set – cropping areas are marked with the dashed lines. a Visible Female
Human. b Visible Ear.

a b

Fig. 2. Image stack of the Visible Ear. The images are symbolized by continuous lines and numbered. The in-
homogeneous original data set (a) was enhanced with additional slices (dashed lines) by copying slides and
squeezing them in. Between slices No. 70 and 335, the distance of the slices is 50 μm, between the other
slices the original distances were 100 μm and are corrected to 50 μm. The enhanced image stack (b) is re-
numbered.

cropped the in-plane size of the data set to 1,225 × 1,225 pixels and converted the file format to the input
format of Vaa3D (i.e. TIFF). The distances between the slices are 50 μm for the part encasing the cochlea and
100 μm for the other portions of the temporal bone. To have equidistant spacing without compression in the
region of the cochlea, duplicate layers were added to regions outside the cochlea so that the whole data set
was enlarged to 944 slices with isotropic voxels (50 × 50 × 50 μm3). To accomplish this, all slices which have
100 μm distances are duplicated and inserted immediately after the original slice reducing slice distance to
50 μm (fig. 2). This final data set with 1,225 × 1,225 × 944 voxels has a total file size of nearly 4 GB (uncom-
pressed), and the volume covers a region of 6.13 × 6.13 × 4.72 cm3. The Visible Ear data set was acquired
from a left human temporal bone.
Importantly, while the order of the data sets from the Visible Ear and the Visible Female Human is
superior to inferior, the slices need to be sorted descending and renumbered ascending otherwise Vaa3D will
not visualize the data set on the correct side of the head (a left ear will be visualized as a right one and vice
versa). For all preprocessing of both data sets (cropping, sorting, adding slices, format conversion), we used
Irfanview with its Advanced Batch Conversion functions.
 ORL 2013;75:46–53 49
DOI: 10.1159/000347083 © 2013 S. Karger AG, Basel
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Kahrs et al.: Freely-Available, True-Color Volume-Rendering Software and
Cryohistology Data Sets for Virtual Exploration of the Temporal Bone Anatomy

Table 1. Comparison of the two data sets used for this study

Original data set Visible Female Human Visible Ear

Color of embedding material Blue Gray

Ear Left and right
Slices inside temporal bone region 230 (1,271 – 1,500)
Original image dimension 4,096 × 3,061 pixels
Original voxel size 0.14 × 0.14 × 0.33 mm3
Extracted regions for visualization 512 × 512 × 230 voxels
7.04 × 7.04 × 7.59 cm3
Crop box coordinates (x, y) (2,201, 1,029) to (2,712, 1,540)
(1,350, 1,085) to (1,861, 1,596)
Automatic sampling factor and resulting None (1 × 1 × 1)
voxel size 0.14 × 0.14 × 0.33 mm3
Left (flipping for right)
All (1 – 605)
3,078 × 1,942 pixels
0.05 × 0.05 × 0.05/0.1 mm3
1,225 × 1,225 × 944 voxels
6.13 × 6.13 × 4.72 cm3
(1,139, 98) to (2,363, 1,322)
0.42 × 0.42 × 0.27
0.12 × 0.12 × 0.18 mm3

After loading an image stack the triplanar view of the data set appears and a 3D rendering (called 3D
view) can be launched. The aforementioned anisotropy factor (e.g. for the Visible Female Human) is needed
to adapt the scaling in the z-direction in this view (setting of z-thickness value). The software has multiple
options for rendering (e.g. choosing threshold of transparency or opacity) and clipping inside the 3D view.
An optimal rendering method for visual exploration is called alpha-value blended, alpha-blending or alpha-
rendering and removes colored regions based on intensity thresholding starting at darker and continuing
towards brighter colors.
While the standard rendering resolution of Vaa3D is 512 × 512 × 256 voxels, larger data sets are auto-
matically downsampled. The visualization of more voxels is possible, but to accomplish such requires making
a change in Vaa3D’s rendering advanced options as well as additional graphics memory on the computer.
Table 1 compares the specifications of both data sets.

Results

When exploring interior structures of both data sets, the Visible Ear appears more natural
concerning color and the rendering has more clarity making some structures easy to identify
and to follow along their course, e.g. facial nerve, carotid artery, inner auditory canal containing
the facial and vestibulocochlear nerves, semicircular canals, ossicles and cochlea. Figure 3
shows the two different volumes used in this study with the triplanar and 3D view using the
program’s alpha-blending. As described above this alpha-rendering method enables one to
show the outer skin surface for the Visible Human (as the skull is embedded in a dark material)
and removes the subarachnoid space for the Visible Ear (as this is one of the darkest struc-
tures in the data set).
Additionally, Vaa3D allows placement of artificial landmarks represented as spherical
markers. They can be placed by the user in one of the views (triplanar or 3D render window)
and will be available in all views. This is helpful for localization of structures which are more
difficult to follow between slices, e.g. it helped us identify the course of the chorda tympani
after initially locating it in one slice following which the spherical markers were visible in 3D
views. Figure 4 shows two exemplary images which can be achieved with Vaa3D and the
Visible Ear data set while clipping the volume inside the 3D rendering view. A fly-through
video is provided as online supplementary material (for all online suppl. material, see www.
karger.com/doi/10.1159/000347083).
 ORL 2013;75:46–53 50
DOI: 10.1159/000347083 © 2013 S. Karger AG, Basel
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Kahrs et al.: Freely-Available, True-Color Volume-Rendering Software and
Cryohistology Data Sets for Virtual Exploration of the Temporal Bone Anatomy

Color version available online

Fig. 3. Both data sets loaded inside Vaa3D and shown in the triplanar and 3D view.
Color version available online

a b

Fig. 4. Two different views inside the Visual Ear data set: more medial (a) and more lateral (b) clipping. Sev-
eral structures are identified and labeled: (1) the ear canal, filled with the embedding material, (2) ossicles,
(3) facial nerve, (4) round window and promontory and (5) semicircular canals.
 ORL 2013;75:46–53 51
DOI: 10.1159/000347083 © 2013 S. Karger AG, Basel
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Kahrs et al.: Freely-Available, True-Color Volume-Rendering Software and
Cryohistology Data Sets for Virtual Exploration of the Temporal Bone Anatomy

Discussion

Computer-aided visualization of histology image stacks from the temporal bone has been
studied for more than 25 years [32]. At present, anatomists and clinicians are encouraged to
mentally visualize complex 3D structure using 2-dimensional images. However, modern
volume and true-color rendering techniques allow usage of such image stacks for 3D visual-
ization and virtual exploration. To date, their use has been limited due to the need for
expensive software as well as the technical knowledge necessary to both set up and run such
software. In this context we present instructions for visualization of the complex anatomy of
the temporal bone using free software and data sets.
In comparison to the free Visible Ear Simulator software [17, 21] or Eaton-Peabody Labo-
ratories 3D Viewer [8], the methods we describe are not limited to specific data sets. Our
intent is to allow establishment of a platform by which users may interact with multiple data
sets both now and in the future. While at present the Visible Ear data set offers the most
complete histological rendering of the temporal bone, future data sets with higher resolution
including microscopic images [33, 34] will likely open up further opportunities for virtual ear
exploration [35]. Ideally, such data sets should be viewable with a single program such that
the user does not need to learn a new program each time a data set becomes available.
While other software packages can carry out similar tasks, Vaa3D is universally available
and offers many options suitable for temporal bone exploration. Clinical CT scans can be
imported into the software as TIFF image stacks as well. The plug-in interface of Vaa3D offers
potential for all kinds of further developments. Haptic feedback as used in other systems [7,
10, 21, 36] is not provided in Vaa3D, but may be doable as a future plug-in. Further enhance-
ment of the software which would be beneficial might include the following: automatic
removal algorithm of pixels representing embedding material [37], automatic reordering of
slices for getting equidistant slices, an automatic method for creating isotropic voxels, inter-
active removal of tissue by virtual ablation or drilling exceeding the current clipping func-
tions, and creating (e.g. tubular) structures from landmarks and visualizing them as surface
renderings directly in the software.
In conclusion, herein we demonstrate a practical solution to understand spatial relation-
ships of the temporal bone using freely available software and data sets. For temporal bone
anatomy, the Visible Ear data set is more complete than the Visible Human. This is likely due
to the increased sampling in the axial direction, better color calibration, fresher specimen,
and more precise image stack alignment. This technique affords an opportunity for virtual
exploration. Ongoing work, coupled at registering clinical CT scans with this data set, may
allow virtual surgery on individual patients prior to actual intervention.

Acknowledgements

This work was supported by Award No. R01DC008408 and R01DC010184 (both to R.F.L.) from the
National Institute on Deafness and Other Communication Disorders (NIDCD, NIH). L.A.K. thanks support by
the German Research Foundation (DFG) within the project KA 2975/2-1. L.A.K. also thanks Jessica Burgner
for help during exploration of the Visible Female Human. The content is solely the responsibility of the
authors and does not necessarily represent the official views of NIDCD, NIH or DFG.

Disclosure Statement

R.F.L. is consultant for Cochlear and is on the Surgical Advisory Boards of MED-EL and Ototronix.
 ORL 2013;75:46–53 52
DOI: 10.1159/000347083 © 2013 S. Karger AG, Basel
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Kahrs et al.: Freely-Available, True-Color Volume-Rendering Software and
Cryohistology Data Sets for Virtual Exploration of the Temporal Bone Anatomy

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